This project deploys a range of structural techniques to examine normal synaptic structure. These approaches have in common their dependence on rapid freezing as well as direct visualization of living brain and both these approaches require direct access to fully viable living brain. Since such preparations are not available, the project in now engaged in exploring various live brain preparations. The most currently active part of this project has been aimed at developing a whole brain preparation that is maintained indefinitely in vitro by vascular perfusion as well as superfusion with artificial CSF. This approach has currently been very successful in maintaining normal structure of the hippocampal and cerebellar cortices for up to 90 minutes of perfusion. Another approach, that has been tried extensively in LN is to develop organotypic brain slice cultures. A comprehensive effort to develop methods for making and maintaining organotypic brain cultures continues (see also Project #ZOl NS 02610-08 LN). It has proven consistently difficult to produce large expanses of mature brain but this preparation is currently being reevaluated. Parts of this project having to do with neuronal development and with synaptic biochemistry have now been established as independent projects (see #s Z01 NS 02871-01 LN and Z01 NS 02872-01 LN).